Human pancreatic secretory trypsin inhibitor stabilizes intestinal mucosa against noxious agents

Pancreatic secretory trypsin inhibitor (PSTI) is a serine protease inhibitor, expressed in gut mucosa, whose function is unclear. We, therefore, examined the effects of PSTI on gut stability and repair. Transgenic mice overexpressing human PSTI within the jejunum (FABPi(-1178 to +28) hPSTI construct) showed no change in baseline morphology or morphometry but reduced indomethacin-induced injury in overexpressing hPSTI region by 42% (P < 0.01). Systemic recombinant hPSTI did not affect baseline morphology or morphometry but truncated injurious effects in prevention and recovery rat models of dextran-sodium-sulfate-induced colitis. In vitro studies showed PSTI stimulated cell migration but not proliferation of human colonic carcinoma HT29 or immortalized mouse colonic YAMC cells. PSTI also induced changes in vectorial ion transport (short-circuit current) when added to basolateral but not apical surfaces of polarized monolayers of Colony-29 cells. Restitution and vectorial ion transport effects of PSTI were dependent on the presence of a functioning epidermal growth factor (EGF) receptor because cells with a disrupted (EGFR(-/-) immortalized cells) or neutralized (EGFR blocking antibodies or tyrosine kinase inhibitor) receptor prevented these effects. PSTI also reduced the cytokine release of lipopolysaccharide-stimulated dendritic cells. We conclude that administration of PSTI may provide a novel method of stabilizing intestinal mucosa against noxious agents and stimulating repair after injury.     

Recirculating CD4 memory T cells mount rapid secondary responses without major contributions from follicular CD4 effectors and B cells

For weeks after primary immunization with thymus-dependent antigens the responding lymph nodes contain effector CD4 T cells in T zones and germinal centers as well as recirculating memory T cells. Conversely, remote nodes, not exposed to antigen, only receive recirculating memory cells. We assessed whether lymph nodes with follicular effector CD4 T cells in addition to recirculating memory CD4 T cells mount a more rapid secondary response than nodes that only contain recirculating memory cells. Also, the extent to which T cell frequency governs accelerated CD4 T cell recall responses was tested. For this, secondary antibody responses to a superantigen, where the frequency of responding T cells is not increased at the time of challenge, were compared with those to conventional protein antigens. With both types of antigens similar accelerated responses were elicited in the node draining the site of primary immunization and in the contralateral node, not previously exposed to antigen. Thus recirculating memory cells are fully capable of mounting accelerated secondary responses, without the assistance of CD4 effector T cells, and accelerated memory responses are not solely dependent on higher T cell frequencies. Accelerated memory CD4 T cell responses were also seen in B cell-deficient mice.     

3,4‐Dihydroxybenzaldehyde lowers ROS generation and protects human red blood cells from arsenic(III) induced oxidative damage

Arsenic (As) is a potent environmental toxicant and chronic exposure to it results in various malignancies in humans. Oxidative stress has been implicated in the etiopathogenesis of As‐induced toxicity. This investigated the protective effect of plant antioxidant 3,4‐dihydroxybenzaldehyde (DHB) on sodium meta‐arsenite (SA), an As‐(III) compound, induced oxidative damage in human red blood cells (RBC). The RBC were first incubated with different concentrations of DHB and then treated with SA at 37°C. Hemolysates were prepared and assayed for various biochemical parameters. Treatment of RBC with SA alone enhanced the generation of reactive oxygen species and increased lipid and protein oxidation. Reduced glutathione levels, total sulfhydryl content and cellular antioxidant power were significantly decreased in SA alone treated RBC, compared to the untreated control cells. This was accompanied by membrane damage, alterations in activities of antioxidant enzymes and deranged glucose metabolism. Incubation of RBC with DHB, prior to treatment with SA, significantly and dose‐dependently attenuated the SA‐induced changes in all these parameters. Scanning electron microscopy of RBC confirmed these biochemical results. Treatment of RBC with SA alone converted the biconcave discoids to echinocytes but the presence of DHB inhibited this conversion and the RBC retained their normal shape. These results show that DHB protects human RBC from SA‐induced oxidative damage, most probably due to its antioxidant character.     

N-hydroxy-pyrroline modification of verapamil exhibits antioxidant protection of the heart against ischemia/reperfusion-induced cardiac dysfunction without compromising its calcium antagonistic activity

Any clinical intervention (e.g., coronary angioplasty, thrombolysis) used to reintroduce blood flow to an ischemic region of the myocardium is accompanied by a complex enzymatic cascade of reactions resulting in severe injury to the heart, termed myocardial ischemia/reperfusion (I/R) injury. In this study, we evaluated the ability of H-3010 (1-hydroxy-2,2,5,5-tetramethyl-2,5-dihydro-1H-pyrrole-3-carboxylic acid (2-(3,4-dimethoxyphenyl)-5-([2-(3,4-dimethoxyphenyl)ethyl]-methylamino)-2-isopropylpentyl)-amide), a pyrroline modification of verapamil (2-(3,4-dimethoxyphenyl)-5-[2-(3,4-dimethoxyphenyl)ethylmethyl-amino]-2-(1-methylethyl)pentanenitrile), to protect the heart against I/R-mediated injury. Isolated perfused rat hearts pretreated with verapamil and H-3010 were subjected to 30 min of global no-flow ischemia followed by 45 min of reperfusion. The recovery (expressed as a percentage of preischemic baseline) in contractile function (left ventricular developed pressure) of hearts subjected to I/R was significantly higher in hearts treated with H-3010 at 5 microM (51.0 +/- 6.4%) as well as at 50 microM (75.1 +/- 7.4%) as compared with verapamil at 5 microM (32.2 +/- 3.7%) or untreated control hearts (18.1 +/- 2.8%). Creatine kinase release was significantly attenuated in hearts treated with H-3010 (45.7 +/- 4.5 U/liter) as compared with untreated controls (131.5 +/- 6.4 U/liter). Similar trends were also observed for lactate dehydrogenase release as well. A marked reduction in percent area of infarction was observed in the H-3010 group (11.7 +/- 1.6%) compared with verapamil (25.1 +/- 2.9%) and control (41.3 +/- 1.9%) groups. Additional in vitro studies showed a marked decrease in reactive oxygen species generation with H-3010. In conclusion, our data clearly demonstrated that the verapamil derivative, H-3010, significantly decreased I/R-induced cardiac dysfunction. This can be attributed to the combined benefits of the pyrroline moiety (antioxidant) and the parent verapamil component (antiarrhythmic) in the protection of the heart from I/R-induced injury.